The major important developments in our recent glycolipid studies are: (1)\Certain glycolipids have been identified as tumor-associated and pre-malignant cell surface markers, and some are characterized as specific differentiation markers. (2)\A close correlation between the ceramide composition and the carbohydrate structure and glycolipid antigenicity has been demonstrated. (3)\Various notions that certain gangliosides, particularly GM3, may play some roles in regulating cell adhesion and cell proliferation have been developed. Defective synthesis of GM3 as had been found in many transformed cells may be related to the loss of cell adhesion and anchorage-independent growth of transformed cells. In view of these developments, we plan to search for new tumor-associated and pre-malignant glycolipid markers and the mechanism of their expression. This project includes systematic fractionation by high-performance liquid chromatography with low-\and high-pressure program, analysis of individual glycolipid and oligosaccharides released by methylation, and by mass spectrometry and charactertization of glycolipids defined by monoclonal antibodies directed to the tumor-distinctive antigen. We will also search for new differentiation markers and chemical and biological characterization. This project will include characterization of glycolipids in teratocarcinoma or myelocyte M1 cells showing differentiation dependence and identification of the stage-specific glycolipid antigen defined by the monoclonal antibodies. Studies will be conducted on the correlation between the ceramide structure, the activity and organization of glycosyltransferases, and on the antigenicity of glycolipids. Finally, the quantity and the organization of GM3 and other specific glycolipids in the adhesion substratum and their changes associated with oncogenic transformation will be studied by cross-linking reagent and with antibodies directed to gangliosides, adhesive proteins, and cytoskeletal components. (A)